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T Ogawa M Hyodo S Ihara M Takekoshi K Minato K Suzuki 《Cell structure and function》1987,12(6):567-574
High-frequency transfection of mouse FM3A cells, grown in suspension, with plasmid pSV2neo DNA was achieved by incubation of the cells with DNA plus polybrene for 6 h followed by an osmotic shock with a hypertonic NaCl solution. When incubated for 20 min at 34 degrees C, FM3A cells showed resistance to the osmolarity change from 0.1 to 9.0% NaCl in the medium. Within this concentration range, 5-7% gave the highest efficiency of transfection. Both linear and circular forms of plasmid DNA produced transformants with equal efficiency. This method was simple, reproducible, and carrier DNA was not required. The efficiency was about 100 times higher than that of the method with DNA-calcium phosphate precipitates. Transformed cells were stable and different numbers of plasmid DNA copies were detected. 相似文献
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Johan Larsbrink Atsushi Izumi Glyn R. Hemsworth Gideon J. Davies Harry Brumer 《The Journal of biological chemistry》2012,287(52):43288-43299
The metabolism of the storage polysaccharides glycogen and starch is of vital importance to organisms from all domains of life. In bacteria, utilization of these α-glucans requires the concerted action of a variety of enzymes, including glycoside hydrolases, glycoside phosphorylases, and transglycosylases. In particular, transglycosylases from glycoside hydrolase family 13 (GH13) and GH77 play well established roles in α-glucan side chain (de)branching, regulation of oligo- and polysaccharide chain length, and formation of cyclic dextrans. Here, we present the biochemical and tertiary structural characterization of a new type of bacterial 1,4-α-glucan 4-α-glucosyltransferase from GH31. Distinct from 1,4-α-glucan 6-α-glucosyltransferases (EC 2.4.1.24) and 4-α-glucanotransferases (EC 2.4.1.25), this enzyme strictly transferred one glucosyl residue from α(1→4)-glucans in disproportionation reactions. Substrate hydrolysis was undetectable for a series of malto-oligosaccharides except maltose for which transglycosylation nonetheless dominated across a range of substrate concentrations. Crystallographic analysis of the enzyme in free, acarbose-complexed, and trapped 5-fluoro-β-glucosyl-enzyme intermediate forms revealed extended substrate interactions across one negative and up to three positive subsites, thus providing structural rationalization for the unique, single monosaccharide transferase activity of the enzyme. 相似文献
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Summary To measure the degree of phr gene induction by DNA-damaging agents, the promoter region was fused to the coding region of the lacZ gene in plasmid pMC1403. The new plasmids were introduced into Escherichia coli cells having different repair capabilities. More efficient induction of phr gene expression was detected in a uvrA
– strain as compared with the wild-type strain. In addition, obvious induction was detected in uvrA
– cells treated by 4-nitroquinoline 1-oxide and mitomycin C. Nalidixic acid, an inhibitor of DNA gyrase, also induced phr gene expression. In contrast, little induced gene expression was noted in UV-irradiated lexA
– and recA
– strains. It is suggested from these results that induction of the phr gene is one of the SOS responses. Possible nucleotide sequences which could be considered to constitute an SOS box were found at the regulator region of the phr gene.Abbreviations
phr
photoreactivation
- UV
ultraviolet light
- 4NQO
4-nitroquinoline 1-oxide
- MMC
mitomycin C
- PRE
photoreactivating enzyme
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E. coli
Escherichia coli 相似文献
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Enzymatic desulfurization of dibenzothiophene by a cell-free system of Rhodococcus erythropolis D-1 总被引:1,自引:0,他引:1
Abstract Pseudomonas syringae cells were exposed to Cu2+ alone or in the precence of acetate, proline or cysteine, at concentrations that reduced free Cu2+ to 1/10 of the total copper. Ligand concentrations (designated as isoeffective) were determined experimentally using a Cu2+ -selective electrode and confirmed by computer calculations using published stability constants. Exposure of P. syringae cells to Cu2+ alone resulted in rapid and pronounced cell death, and binding of most of the copper in solution. The addition of acetate, proline or cysteine, a few minutes after Cu2+ treatment, resulted in a significant reduction in cell death, and in the amount of copper bound to the cells. For short exposures to Cu2+ , cysteine was more effective than acetate or proline, but after 60 min of treatment, similar results were observed with these ligands. The addition of ligands before Cu2+ resulted in even more reduced copper toxicity. The results showed that, at isoeffective concentrations, weak and moderate copper-ligands can effectively antagonize copper toxicity, and that this protective effect does not require previously equilibrated copper-ligand solutions and is not very dependent of the nature of the ligand. 相似文献
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Takashi Yokochi Izumi Nakashima Nobuo Kato Junpei Asai Soichi Iijima 《Microbiology and immunology》1980,24(10):933-944
The sequence of histological changes in the regional lymph node and other lymphoid organs of mice injected with the capsular polysaccharide of Klebsiella pneumoniae (CPS-K) or bacterial lipopolysaccharide (LPS) was followed. Injection of CPS-K, but not LPS, induced the following characteristic histological changes in the regional lymph node. In the early stage there was a marked decrease in the number of small lymphocytes, accompanied by the appearance of scattered fragmented nuclei and infiltration of polymorphonuclear neutrophilic leukocytes, and in the late stage there was marked proliferation of macrophage-like cells and pyroninophilic cells. Histological changes in the thymus and spleen and changes in cell populations in the bone marrow and peripheral blood after CPS-K injection were essentially the same as after LPS injection. Since CPS-K has a much stronger adjuvant action on antibody response than does LPS, it is suggested that the characteristic histological changes in the regional lymph node after injection of CPS-K are closely related to its extraordinarily strong adjuvant action. 相似文献
10.
Sakuya Nakamura Shinya Hagihara Masanori Izumi 《Biochimica et Biophysica Acta (BBA)/General Subjects》2021,1865(8):129916
Mitochondria play a central role in primary metabolism in plants as well as in heterotrophic eukaryotes. Plants must control the quality and number of mitochondria in response to a changing environment, across cell types and developmental stages. Mitophagy is defined as the degradation of mitochondria by autophagy, an evolutionarily conserved system for the removal and recycling of intracellular components. Recent studies have highlighted the importance of mitophagy in plant stress responses. This review article summarizes our current knowledge of plant mitophagy and discusses the underlying mechanisms. In plants, chloroplasts cooperate with mitochondria for energy production, and autophagy also targets chloroplasts through a process known as chlorophagy. Advances in plant autophagy studies now allow a comparative analysis of the autophagic turnover of mitochondria and chloroplasts, via the selective degradation of their soluble proteins, fragments, or entire organelles. 相似文献